The specific aim of this proposal is to define the manner in which intracellular T3 concentrations are regulated in different tissues. There are two potential sources of tissue T3; from plasma, T3(T3), or from local conversion of T4 to T3 within the tissue, T3(T4). In the CNS and pituitary, T3(T4) is predominant whereas in liver and kidney, T3(T3) comprises the bulk of intracellular T3. The presence of tissue T3(T4) cannot be predicted from the plasma T3. By using physiological and biochemical techniques, we will determine the reasons for the differences between various tissues with respect to tissue T3 sources. We will evaluate the lung, skeletal muscle, small intestine and spleen to determine the sources of intracellular T3 and confirm these estimates by RIA. We will study the tissue kinetics of tracer T3, T4, and T3(T4) in the cortex and compare these results to those in liver and kidney of euthyroid rats. Since the activity of the enzyme converting T4 to T3 in the brain responds in opposite directions to changes in thyroid status from that in the liver, the effects of these physiological perturbations on intracellular T3 and T4 kinetics will also be evaluated. The iodine-deficient rat will also be examined since T4 to T3 conversion in brain should be increased in this animal. Effects of thyroid status on serum and cytosolic thyroid hormone binding proteins will be documented. The results obtained in intact animals will be explored under more carefully controlled conditions in tissue culture. Embryonic brain tissue from fetal rats will be used to study thyroid hormone metabolism at a cellular level and to determine the effects of thyroid status on the relative contributions of T3(T4) and T3(T3) in the in vitro system. Surviving pituitary cells in culture will also be evaluated especially to compare thyrotrophs with other pituitary cells. These studies should provide insights as to tissue-specific mechanisms for regulation of intracellular T3. This knowledge is important for understanding the proper application and limitations of thyroid function tests. It has broad implications for the diagnosis, treatment and monitoring of patients with thyroid diseases as well as for evaluation of thyroid status in patients with acute illnesses where T4 and T3 conversion is impaired.